The Experts below are selected from a list of 1989 Experts worldwide ranked by ideXlab platform
Stanley M Spinola - One of the best experts on this subject based on the ideXlab platform.
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Determination of an Interaction Network between an Extracellular Bacterial Pathogen and the Human Host
'American Society for Microbiology', 2019Co-Authors: Brad Griesenauer, Diane M. Janowicz, Kate R Fortney, Tuan M. Tran, Paula Johnson, Hongyu Gao, Stephen Barnes, Landon S. Wilson, Yunlong Liu, Stanley M SpinolaAbstract:Dual RNA sequencing (RNA-seq) offers the promise of determining an interactome at a transcriptional level between a bacterium and the host but has yet to be done on any bacterial infection in human tissue. We performed dual RNA-seq and metabolomics analyses on wounded and infected sites following experimental infection of the arm with H. ducreyi. Our results suggest that H. ducreyi survives in an abscess by utilizing l-ascorbate as an alternative carbon source, possibly taking advantage of host ascorbic acid recycling, and that H. ducreyi also adapts by upregulating genes involved in anaerobic metabolism and inorganic ion and nutrient transport. To our knowledge, this is the first description of an interaction network between a bacterium and the human host at a site of infection.A major gap in understanding infectious diseases is the lack of information about molecular interaction networks between pathogens and the human host. Haemophilus ducreyi causes the genital ulcer disease chancroid in adults and is a leading cause of cutaneous ulcers in children in the tropics. We developed a model in which human volunteers are infected on the upper arm with H. ducreyi until they develop Pustules. To define the H. ducreyi and human interactome, we determined bacterial and host transcriptomic and host metabolomic changes in Pustules. We found that in vivoH. ducreyi transcripts were distinct from those in the inocula, as were host transcripts in Pustule and wounded control sites. Many of the upregulated H. ducreyi genes were found to be involved in ascorbic acid and anaerobic metabolism and inorganic ion/nutrient transport. The top 20 significantly expressed human pathways showed that all were involved in immune responses. We generated a bipartite network for interactions between host and bacterial gene transcription; multiple positively correlated networks contained H. ducreyi genes involved in anaerobic metabolism and host genes involved with the immune response. Metabolomic studies showed that Pustule and wounded samples had different metabolite compositions; the top ion pathway involved ascorbate and aldarate metabolism, which correlated with the H. ducreyi transcriptional response and upregulation of host genes involved in ascorbic acid recycling. These data show that an interactome exists between H. ducreyi and the human host and suggest that H. ducreyi exploits the metabolic niche created by the host immune response
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host polymorphisms in tlr9 and il10 are associated with the outcomes of experimental haemophilus ducreyi infection in human volunteers
The Journal of Infectious Diseases, 2016Co-Authors: M Singer, Servaas A Morre, Sander Ouburg, Stanley M SpinolaAbstract:BACKGROUND In humans inoculated with Haemophilus ducreyi, there are host effects on the possible clinical outcomes-Pustule formation versus spontaneous resolution of infection. However, the immunogenetic factors that influence these outcomes are unknown. Here we examined the role of 14 single-nucleotide polymorphisms (SNPs) in 7 selected pathogen-recognition pathways and cytokine genes on the gradated outcomes of experimental infection. METHODS DNAs from 105 volunteers infected with H. ducreyi at 3 sites were genotyped for SNPs, using real-time polymerase chain reaction. The participants were classified into 2 cohorts, by race, and into 4 groups, based on whether they formed 0, 1, 2, or 3 Pustules. χ(2) tests for trend and logistic regression analyses were performed on the data. RESULTS In European Americans, the most significant findings were a protective association of the TLR9 +2848 GG genotype and a risk-enhancing association of the TLR9 TA haplotype with Pustule formation; logistic regression showed a trend toward protection for the TLR9 +2848 GG genotype. In African Americans, logistic regression showed a protective effect for the IL10 -2849 AA genotype and a risk-enhancing effect for the IL10 AAC haplotype. CONCLUSIONS Variations in TLR9 and IL10 are associated with the outcome of H. ducreyi infection.
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role played by cd4 foxp3 regulatory t cells in suppression of host responses to haemophilus ducreyi during experimental infection of human volunteers
The Journal of Infectious Diseases, 2010Co-Authors: Klara Tennerracz, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, Paul Racz, Stanley M SpinolaAbstract:Haemophilus ducreyi is the causative agent of the sexually transmitted disease chancroid, a genital ulcer disease that facilitates the acquisition and transmission of HIV-1 [1]. In order to understand the immunopathogenesis of H. ducreyi infection, we developed a human challenge model, in which the skin of the upper arm of healthy adult volunteers is inoculated with the H. ducreyi strain 35000HP (HP, human passaged) or its derivatives [2]. After inoculation, papules form within 24 hours and either spontaneously resolve or evolve into Pustules within 2 to 5 days. Experimental Pustules and natural ulcers are identical histologically and signify immunological failure. Both innate and adaptive immune cells including neutrophils, macrophages, myeloid dendritic cells (DC), NK cells and memory/effector T cells are recruited to experimental Pustules and natural ulcers [3–5]. In Pustules, neutrophils coalesce to form an epidermal abscess, and macrophages form a collar at the base of the abscess. Below the collar, there is a dermal infiltrate of T cells, NK cells and macrophages. Despite this response, H. ducreyi replicates and persists at infected sites. In Pustules and natural ulcers, the bacterium colocalizes with neutrophils and macrophages and remains extracellular [6, 7]. Thus, evasion of phagocytosis is a major mechanism of bacterial survival in experimental and natural infection. Regulatory T (Treg) cells actively suppress the function of the adaptive and innate immune systems [8]. Treg cells are essential for maintaining self-tolerance and immune homeostasis. Treg cells use cell-contact inhibition or soluble factors to control collateral tissue damage mediated by immune responses. However, they also prevent sterilizing antimicrobial immunity and promote pathogen persistence during infection. Two major types of Treg cells have been described based on their origin of generation [9]. Naturally occurring Treg (nTreg) cells are generated in the thymus and express CD25+ and the transcription factor forkhead box P3 (FOXP3), which is critical for their development and function. Treg cells can also be converted from mature CD4+CD25− T cells in peripheral tissues under immunosuppressive conditions, such as exposure to IL-10, transforming growth factor-β (TGF-β) or indoleamine 2,3-dioxygenase (IDO) made by APC [10–14]. These inducible Treg (iTreg) cells are classified as IL-10-producing Tr1 cells, TGF-β-producing Th3 cells and inducible FOXP3+ Treg cells. After infection with pathogens, Treg cells accumulate at infected sites through recruitment, retention, proliferation and/or conversion [15]. Clinical and laboratory data suggest that Treg cells may be involved in the formation of H. ducreyi-induced Pustules. Approximately 16% of volunteers develop hypertrophic scars at H. ducreyi-infected sites that are biopsied [2]. Hypertrophic scar formation is associated with production of TGF-β, which promotes the development of Treg cells. Upon exposure to H. ducreyi, monocyte-derived DC from Pustule formers upregulate transcripts of markers that foster Treg development [16]. Indeed, some H. ducreyi-specific T cell lines and clones isolated from Pustules have characteristics of Treg cells in that they produce IL-10 and IFN-γ [17]. Here, we tested the hypothesis that Treg cells accumulate in H. ducreyi-infected Pustules. We found that CD4+FOXP3+ T cells were enriched in experimental Pustules and were mainly located at the base of Pustules. Most of the CD4+FOXP3+ T cells expressed phenotypic markers characteristic of circulating CD4+FOXP3+ Treg cells. We also showed that the CD4+FOXP3+ T cells were unable to produce effector cytokines and that depletion of CD4+CD25+ T cells from PBMCs of persons who formed Pustules and uninfected donors increased proliferative response of CD4 T cells to H. ducreyi.
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experimental infection of human volunteers with haemophilus ducreyi fifteen years of clinical data and experience
The Journal of Infectious Diseases, 2009Co-Authors: Diane M. Janowicz, Barry P. Katz, Susan Ofner, Stanley M SpinolaAbstract:Haemophilus ducreyi causes chancroid, which facilitates transmission of human immunodeficiency virus type 1. To better understand the biology of H. ducreyi, we developed a human inoculation model. In the present article, we describe clinical outcomes for 267 volunteers who were infected with H. ducreyi. There was a relationship between papule formation and estimated delivered dose. The outcome (either Pustule formation or resolution) of infected sites for a given subject was not independent; the most important determinants of Pustule formation were sex and host effects. When 41 subjects were infected a second time, their outcomes segregated toward their initial outcome, confirming the host effect. Subjects with Pustules developed local symptoms that required withdrawal from the study after a mean of 8.6 days. There were 191 volunteers who had tissue biopsy performed, 173 of whom were available for follow-up analysis; 28 (16.2%) of these developed hypertrophic scars, but the model was otherwise safe. Mutant-parent trials confirmed key features in H. ducreyi pathogenesis, and the model has provided an opportunity to study differential human susceptibility to a bacterial infection.
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evaluation of the repertoire of the tonb dependent receptors of haemophilus ducreyi for their role in virulence in humans
The Journal of Infectious Diseases, 2008Co-Authors: Isabelle Leduc, Kate R Fortney, Stanley M Spinola, Barry P. Katz, Keith E Banks, Kristine B Patterson, Steve D Billings, Christopher ElkinsAbstract:Haemophilus ducreyi contains 3 TonB-dependent receptors: the hemoglobin receptor HgbA, which is required for virulence in humans; the heme receptor TdhA; and an uncharacterized conserved hypothetical protein TdX (HD0646). A double tdX/tdhA mutant (FX527) was constructed on the background of a human-passaged variant of strain 35000 (35000HP). Six volunteers were infected with 35000HP at 3 sites on one arm and with FX527 at 3 sites on the other. The Pustule formation rate was 55.6% (95% confidence interval [CI], 35.7%-75.4%) at 18 parent-strain sites and 44.4% (95% CI, 15.0%-73.9%) at 18 mutant-strain sites (P = .51). Similar amounts of 35000HP and FX527 were recovered from Pustules in semiquantitative culture. Thus, TdX and TdhA are not necessary for virulence, whereas HgbA is both necessary and sufficient for virulence in humans. The data suggest that hemoglobin is the sole source of heme/iron used by H. ducreyi in vivo and has implications for the potential of HgbA as a vaccine.
Barry P. Katz - One of the best experts on this subject based on the ideXlab platform.
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The Human Skin Microbiome Associates with the Outcome of and Is Influenced by Bacterial Infection
Mbio, 2015Co-Authors: Julia J. Van Rensburg, Sheila Ellinger, Beth Zwickl, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, David E NelsonAbstract:ABSTRACT The influence of the skin microbiota on host susceptibility to infectious agents is largely unexplored. The skin harbors diverse bacterial species that may promote or antagonize the growth of an invading pathogen. We developed a human infection model for Haemophilus ducreyi in which human volunteers are inoculated on the upper arm. After inoculation, papules form and either spontaneously resolve or progress to Pustules. To examine the role of the skin microbiota in the outcome of H. ducreyi infection, we analyzed the microbiomes of four dose-matched pairs of “resolvers” and “Pustule formers” whose inoculation sites were swabbed at multiple time points. Bacteria present on the skin were identified by amplification and pyrosequencing of 16S rRNA genes. Nonmetric multidimensional scaling (NMDS) using Bray-Curtis dissimilarity between the preinfection microbiomes of infected sites showed that sites from the same volunteer clustered together and that Pustule formers segregated from resolvers ( P = 0.001, permutational multivariate analysis of variance [PERMANOVA]), suggesting that the preinfection microbiomes were associated with outcome. NMDS using Bray-Curtis dissimilarity of the endpoint samples showed that the Pustule sites clustered together and were significantly different than the resolved sites ( P = 0.001, PERMANOVA), suggesting that the microbiomes at the endpoint differed between the two groups. In addition to H. ducreyi, Pustule-forming sites had a greater abundance of Proteobacteria , Bacteroidetes , Micrococcus , Corynebacterium , Paracoccus , and Staphylococcus species, whereas resolved sites had higher levels of Actinobacteria and Propionibacterium species. These results suggest that at baseline, resolvers and Pustule formers have distinct skin bacterial communities which change in response to infection and the resultant immune response. IMPORTANCE Human skin is home to a diverse community of microorganisms, collectively known as the skin microbiome. Some resident bacteria are thought to protect the skin from infection by outcompeting pathogens for resources or by priming the immune system9s response to invaders. However, the influence of the skin microbiome on the susceptibility to or protection from infection has not been prospectively evaluated in humans. We characterized the skin microbiome before, during, and after experimental inoculation of the arm with Haemophilus ducreyi in matched volunteers who subsequently resolved the infection or formed abscesses. Our results suggest that the preinfection microbiomes of Pustule formers and resolvers have distinct community structures which change in response to the progression of H. ducreyi infection to abscess formation.
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role played by cd4 foxp3 regulatory t cells in suppression of host responses to haemophilus ducreyi during experimental infection of human volunteers
The Journal of Infectious Diseases, 2010Co-Authors: Klara Tennerracz, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, Paul Racz, Stanley M SpinolaAbstract:Haemophilus ducreyi is the causative agent of the sexually transmitted disease chancroid, a genital ulcer disease that facilitates the acquisition and transmission of HIV-1 [1]. In order to understand the immunopathogenesis of H. ducreyi infection, we developed a human challenge model, in which the skin of the upper arm of healthy adult volunteers is inoculated with the H. ducreyi strain 35000HP (HP, human passaged) or its derivatives [2]. After inoculation, papules form within 24 hours and either spontaneously resolve or evolve into Pustules within 2 to 5 days. Experimental Pustules and natural ulcers are identical histologically and signify immunological failure. Both innate and adaptive immune cells including neutrophils, macrophages, myeloid dendritic cells (DC), NK cells and memory/effector T cells are recruited to experimental Pustules and natural ulcers [3–5]. In Pustules, neutrophils coalesce to form an epidermal abscess, and macrophages form a collar at the base of the abscess. Below the collar, there is a dermal infiltrate of T cells, NK cells and macrophages. Despite this response, H. ducreyi replicates and persists at infected sites. In Pustules and natural ulcers, the bacterium colocalizes with neutrophils and macrophages and remains extracellular [6, 7]. Thus, evasion of phagocytosis is a major mechanism of bacterial survival in experimental and natural infection. Regulatory T (Treg) cells actively suppress the function of the adaptive and innate immune systems [8]. Treg cells are essential for maintaining self-tolerance and immune homeostasis. Treg cells use cell-contact inhibition or soluble factors to control collateral tissue damage mediated by immune responses. However, they also prevent sterilizing antimicrobial immunity and promote pathogen persistence during infection. Two major types of Treg cells have been described based on their origin of generation [9]. Naturally occurring Treg (nTreg) cells are generated in the thymus and express CD25+ and the transcription factor forkhead box P3 (FOXP3), which is critical for their development and function. Treg cells can also be converted from mature CD4+CD25− T cells in peripheral tissues under immunosuppressive conditions, such as exposure to IL-10, transforming growth factor-β (TGF-β) or indoleamine 2,3-dioxygenase (IDO) made by APC [10–14]. These inducible Treg (iTreg) cells are classified as IL-10-producing Tr1 cells, TGF-β-producing Th3 cells and inducible FOXP3+ Treg cells. After infection with pathogens, Treg cells accumulate at infected sites through recruitment, retention, proliferation and/or conversion [15]. Clinical and laboratory data suggest that Treg cells may be involved in the formation of H. ducreyi-induced Pustules. Approximately 16% of volunteers develop hypertrophic scars at H. ducreyi-infected sites that are biopsied [2]. Hypertrophic scar formation is associated with production of TGF-β, which promotes the development of Treg cells. Upon exposure to H. ducreyi, monocyte-derived DC from Pustule formers upregulate transcripts of markers that foster Treg development [16]. Indeed, some H. ducreyi-specific T cell lines and clones isolated from Pustules have characteristics of Treg cells in that they produce IL-10 and IFN-γ [17]. Here, we tested the hypothesis that Treg cells accumulate in H. ducreyi-infected Pustules. We found that CD4+FOXP3+ T cells were enriched in experimental Pustules and were mainly located at the base of Pustules. Most of the CD4+FOXP3+ T cells expressed phenotypic markers characteristic of circulating CD4+FOXP3+ Treg cells. We also showed that the CD4+FOXP3+ T cells were unable to produce effector cytokines and that depletion of CD4+CD25+ T cells from PBMCs of persons who formed Pustules and uninfected donors increased proliferative response of CD4 T cells to H. ducreyi.
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experimental infection of human volunteers with haemophilus ducreyi fifteen years of clinical data and experience
The Journal of Infectious Diseases, 2009Co-Authors: Diane M. Janowicz, Barry P. Katz, Susan Ofner, Stanley M SpinolaAbstract:Haemophilus ducreyi causes chancroid, which facilitates transmission of human immunodeficiency virus type 1. To better understand the biology of H. ducreyi, we developed a human inoculation model. In the present article, we describe clinical outcomes for 267 volunteers who were infected with H. ducreyi. There was a relationship between papule formation and estimated delivered dose. The outcome (either Pustule formation or resolution) of infected sites for a given subject was not independent; the most important determinants of Pustule formation were sex and host effects. When 41 subjects were infected a second time, their outcomes segregated toward their initial outcome, confirming the host effect. Subjects with Pustules developed local symptoms that required withdrawal from the study after a mean of 8.6 days. There were 191 volunteers who had tissue biopsy performed, 173 of whom were available for follow-up analysis; 28 (16.2%) of these developed hypertrophic scars, but the model was otherwise safe. Mutant-parent trials confirmed key features in H. ducreyi pathogenesis, and the model has provided an opportunity to study differential human susceptibility to a bacterial infection.
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evaluation of the repertoire of the tonb dependent receptors of haemophilus ducreyi for their role in virulence in humans
The Journal of Infectious Diseases, 2008Co-Authors: Isabelle Leduc, Kate R Fortney, Stanley M Spinola, Barry P. Katz, Keith E Banks, Kristine B Patterson, Steve D Billings, Christopher ElkinsAbstract:Haemophilus ducreyi contains 3 TonB-dependent receptors: the hemoglobin receptor HgbA, which is required for virulence in humans; the heme receptor TdhA; and an uncharacterized conserved hypothetical protein TdX (HD0646). A double tdX/tdhA mutant (FX527) was constructed on the background of a human-passaged variant of strain 35000 (35000HP). Six volunteers were infected with 35000HP at 3 sites on one arm and with FX527 at 3 sites on the other. The Pustule formation rate was 55.6% (95% confidence interval [CI], 35.7%-75.4%) at 18 parent-strain sites and 44.4% (95% CI, 15.0%-73.9%) at 18 mutant-strain sites (P = .51). Similar amounts of 35000HP and FX527 were recovered from Pustules in semiquantitative culture. Thus, TdX and TdhA are not necessary for virulence, whereas HgbA is both necessary and sufficient for virulence in humans. The data suggest that hemoglobin is the sole source of heme/iron used by H. ducreyi in vivo and has implications for the potential of HgbA as a vaccine.
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a dlta mutant of haemophilus ducreyi is partially attenuated in its ability to cause Pustules in human volunteers
Infection and Immunity, 2006Co-Authors: Diane M. Janowicz, Kate R Fortney, Barry P. Katz, Isabelle Leduc, Christopher Elkins, Stanley M SpinolaAbstract:Haemophilus ducreyi produces two outer membrane proteins, called DltA (H. ducreyi lectin A) and DsrA (H. ducreyi serum resistance A), that contribute to the ability of the organism to evade complement-mediated serum killing. In contrast to their isogenic parent strain, 35000HP, the DsrA mutant FX517 exhibits 0% survival in 50% normal human serum and the DltA mutant FX533 exhibits 23% survival. Compared to 35000HP, FX517 does not cause Pustule formation in human volunteers. To test whether DltA was required for virulence in humans, seven volunteers were experimentally infected with 35000HP and FX533. Four subjects were inoculated with fixed doses of 35000HP (101 CFU or 130 CFU) at three sites on one arm and escalating doses of FX533 (range, 46 CFU to 915 CFU) at three sites on the other arm. Pustules only developed at mutant-injected sites at doses nearly twofold higher than that of the parent, suggesting that FX533 was partially attenuated. Three subjects were inoculated with similar doses of the parent (67 CFU) and mutant (104 CFU) at three sites. Pustules formed at five of nine parent sites and one of nine mutant sites. Overall, the papule and Pustule formation rates for 35000HP and FX533 were similar for the trial. However, for the five subjects who received similar doses of the parent and mutant, Pustules developed at 7 of 15 sites (46.7%; 95% confidence interval [CI], 16.9% to 76.5%) inoculated with the parent and at 1 of 15 (6.7%; 95% CI, 0.1% to 18.4%) sites inoculated with the mutant (P = 0.043). We concluded that the DltA mutant was attenuated in its ability to cause disease at doses similar to that of the parent.
Kate R Fortney - One of the best experts on this subject based on the ideXlab platform.
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Determination of an Interaction Network between an Extracellular Bacterial Pathogen and the Human Host
'American Society for Microbiology', 2019Co-Authors: Brad Griesenauer, Diane M. Janowicz, Kate R Fortney, Tuan M. Tran, Paula Johnson, Hongyu Gao, Stephen Barnes, Landon S. Wilson, Yunlong Liu, Stanley M SpinolaAbstract:Dual RNA sequencing (RNA-seq) offers the promise of determining an interactome at a transcriptional level between a bacterium and the host but has yet to be done on any bacterial infection in human tissue. We performed dual RNA-seq and metabolomics analyses on wounded and infected sites following experimental infection of the arm with H. ducreyi. Our results suggest that H. ducreyi survives in an abscess by utilizing l-ascorbate as an alternative carbon source, possibly taking advantage of host ascorbic acid recycling, and that H. ducreyi also adapts by upregulating genes involved in anaerobic metabolism and inorganic ion and nutrient transport. To our knowledge, this is the first description of an interaction network between a bacterium and the human host at a site of infection.A major gap in understanding infectious diseases is the lack of information about molecular interaction networks between pathogens and the human host. Haemophilus ducreyi causes the genital ulcer disease chancroid in adults and is a leading cause of cutaneous ulcers in children in the tropics. We developed a model in which human volunteers are infected on the upper arm with H. ducreyi until they develop Pustules. To define the H. ducreyi and human interactome, we determined bacterial and host transcriptomic and host metabolomic changes in Pustules. We found that in vivoH. ducreyi transcripts were distinct from those in the inocula, as were host transcripts in Pustule and wounded control sites. Many of the upregulated H. ducreyi genes were found to be involved in ascorbic acid and anaerobic metabolism and inorganic ion/nutrient transport. The top 20 significantly expressed human pathways showed that all were involved in immune responses. We generated a bipartite network for interactions between host and bacterial gene transcription; multiple positively correlated networks contained H. ducreyi genes involved in anaerobic metabolism and host genes involved with the immune response. Metabolomic studies showed that Pustule and wounded samples had different metabolite compositions; the top ion pathway involved ascorbate and aldarate metabolism, which correlated with the H. ducreyi transcriptional response and upregulation of host genes involved in ascorbic acid recycling. These data show that an interactome exists between H. ducreyi and the human host and suggest that H. ducreyi exploits the metabolic niche created by the host immune response
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The Human Skin Microbiome Associates with the Outcome of and Is Influenced by Bacterial Infection
Mbio, 2015Co-Authors: Julia J. Van Rensburg, Sheila Ellinger, Beth Zwickl, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, David E NelsonAbstract:ABSTRACT The influence of the skin microbiota on host susceptibility to infectious agents is largely unexplored. The skin harbors diverse bacterial species that may promote or antagonize the growth of an invading pathogen. We developed a human infection model for Haemophilus ducreyi in which human volunteers are inoculated on the upper arm. After inoculation, papules form and either spontaneously resolve or progress to Pustules. To examine the role of the skin microbiota in the outcome of H. ducreyi infection, we analyzed the microbiomes of four dose-matched pairs of “resolvers” and “Pustule formers” whose inoculation sites were swabbed at multiple time points. Bacteria present on the skin were identified by amplification and pyrosequencing of 16S rRNA genes. Nonmetric multidimensional scaling (NMDS) using Bray-Curtis dissimilarity between the preinfection microbiomes of infected sites showed that sites from the same volunteer clustered together and that Pustule formers segregated from resolvers ( P = 0.001, permutational multivariate analysis of variance [PERMANOVA]), suggesting that the preinfection microbiomes were associated with outcome. NMDS using Bray-Curtis dissimilarity of the endpoint samples showed that the Pustule sites clustered together and were significantly different than the resolved sites ( P = 0.001, PERMANOVA), suggesting that the microbiomes at the endpoint differed between the two groups. In addition to H. ducreyi, Pustule-forming sites had a greater abundance of Proteobacteria , Bacteroidetes , Micrococcus , Corynebacterium , Paracoccus , and Staphylococcus species, whereas resolved sites had higher levels of Actinobacteria and Propionibacterium species. These results suggest that at baseline, resolvers and Pustule formers have distinct skin bacterial communities which change in response to infection and the resultant immune response. IMPORTANCE Human skin is home to a diverse community of microorganisms, collectively known as the skin microbiome. Some resident bacteria are thought to protect the skin from infection by outcompeting pathogens for resources or by priming the immune system9s response to invaders. However, the influence of the skin microbiome on the susceptibility to or protection from infection has not been prospectively evaluated in humans. We characterized the skin microbiome before, during, and after experimental inoculation of the arm with Haemophilus ducreyi in matched volunteers who subsequently resolved the infection or formed abscesses. Our results suggest that the preinfection microbiomes of Pustule formers and resolvers have distinct community structures which change in response to the progression of H. ducreyi infection to abscess formation.
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role played by cd4 foxp3 regulatory t cells in suppression of host responses to haemophilus ducreyi during experimental infection of human volunteers
The Journal of Infectious Diseases, 2010Co-Authors: Klara Tennerracz, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, Paul Racz, Stanley M SpinolaAbstract:Haemophilus ducreyi is the causative agent of the sexually transmitted disease chancroid, a genital ulcer disease that facilitates the acquisition and transmission of HIV-1 [1]. In order to understand the immunopathogenesis of H. ducreyi infection, we developed a human challenge model, in which the skin of the upper arm of healthy adult volunteers is inoculated with the H. ducreyi strain 35000HP (HP, human passaged) or its derivatives [2]. After inoculation, papules form within 24 hours and either spontaneously resolve or evolve into Pustules within 2 to 5 days. Experimental Pustules and natural ulcers are identical histologically and signify immunological failure. Both innate and adaptive immune cells including neutrophils, macrophages, myeloid dendritic cells (DC), NK cells and memory/effector T cells are recruited to experimental Pustules and natural ulcers [3–5]. In Pustules, neutrophils coalesce to form an epidermal abscess, and macrophages form a collar at the base of the abscess. Below the collar, there is a dermal infiltrate of T cells, NK cells and macrophages. Despite this response, H. ducreyi replicates and persists at infected sites. In Pustules and natural ulcers, the bacterium colocalizes with neutrophils and macrophages and remains extracellular [6, 7]. Thus, evasion of phagocytosis is a major mechanism of bacterial survival in experimental and natural infection. Regulatory T (Treg) cells actively suppress the function of the adaptive and innate immune systems [8]. Treg cells are essential for maintaining self-tolerance and immune homeostasis. Treg cells use cell-contact inhibition or soluble factors to control collateral tissue damage mediated by immune responses. However, they also prevent sterilizing antimicrobial immunity and promote pathogen persistence during infection. Two major types of Treg cells have been described based on their origin of generation [9]. Naturally occurring Treg (nTreg) cells are generated in the thymus and express CD25+ and the transcription factor forkhead box P3 (FOXP3), which is critical for their development and function. Treg cells can also be converted from mature CD4+CD25− T cells in peripheral tissues under immunosuppressive conditions, such as exposure to IL-10, transforming growth factor-β (TGF-β) or indoleamine 2,3-dioxygenase (IDO) made by APC [10–14]. These inducible Treg (iTreg) cells are classified as IL-10-producing Tr1 cells, TGF-β-producing Th3 cells and inducible FOXP3+ Treg cells. After infection with pathogens, Treg cells accumulate at infected sites through recruitment, retention, proliferation and/or conversion [15]. Clinical and laboratory data suggest that Treg cells may be involved in the formation of H. ducreyi-induced Pustules. Approximately 16% of volunteers develop hypertrophic scars at H. ducreyi-infected sites that are biopsied [2]. Hypertrophic scar formation is associated with production of TGF-β, which promotes the development of Treg cells. Upon exposure to H. ducreyi, monocyte-derived DC from Pustule formers upregulate transcripts of markers that foster Treg development [16]. Indeed, some H. ducreyi-specific T cell lines and clones isolated from Pustules have characteristics of Treg cells in that they produce IL-10 and IFN-γ [17]. Here, we tested the hypothesis that Treg cells accumulate in H. ducreyi-infected Pustules. We found that CD4+FOXP3+ T cells were enriched in experimental Pustules and were mainly located at the base of Pustules. Most of the CD4+FOXP3+ T cells expressed phenotypic markers characteristic of circulating CD4+FOXP3+ Treg cells. We also showed that the CD4+FOXP3+ T cells were unable to produce effector cytokines and that depletion of CD4+CD25+ T cells from PBMCs of persons who formed Pustules and uninfected donors increased proliferative response of CD4 T cells to H. ducreyi.
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evaluation of the repertoire of the tonb dependent receptors of haemophilus ducreyi for their role in virulence in humans
The Journal of Infectious Diseases, 2008Co-Authors: Isabelle Leduc, Kate R Fortney, Stanley M Spinola, Barry P. Katz, Keith E Banks, Kristine B Patterson, Steve D Billings, Christopher ElkinsAbstract:Haemophilus ducreyi contains 3 TonB-dependent receptors: the hemoglobin receptor HgbA, which is required for virulence in humans; the heme receptor TdhA; and an uncharacterized conserved hypothetical protein TdX (HD0646). A double tdX/tdhA mutant (FX527) was constructed on the background of a human-passaged variant of strain 35000 (35000HP). Six volunteers were infected with 35000HP at 3 sites on one arm and with FX527 at 3 sites on the other. The Pustule formation rate was 55.6% (95% confidence interval [CI], 35.7%-75.4%) at 18 parent-strain sites and 44.4% (95% CI, 15.0%-73.9%) at 18 mutant-strain sites (P = .51). Similar amounts of 35000HP and FX527 were recovered from Pustules in semiquantitative culture. Thus, TdX and TdhA are not necessary for virulence, whereas HgbA is both necessary and sufficient for virulence in humans. The data suggest that hemoglobin is the sole source of heme/iron used by H. ducreyi in vivo and has implications for the potential of HgbA as a vaccine.
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identification of haemophilus ducreyi genes expressed during human infection
Microbiology, 2008Co-Authors: Margaret E Bauer, Diane M. Janowicz, Kate R Fortney, Robert S Munson, Alistair Harrison, Stanley M SpinolaAbstract:To identify Haemophilus ducreyi transcripts that are expressed during human infection, we used selective capture of transcribed sequences (SCOTS) with RNA isolated from Pustules obtained from three volunteers infected with H. ducreyi, and with RNA isolated from broth-grown bacteria used to infect volunteers. With SCOTS, competitive hybridization of tissue-derived and broth-derived sequences identifies genes that may be preferentially expressed in vivo. Among the three tissue specimens, we identified 531 genes expressed in vivo. Southern blot analysis of 60 genes from each tissue showed that 87 % of the identified genes hybridized better with cDNA derived from tissue specimens than with cDNA derived from broth-grown bacteria. RT-PCR on nine additional Pustules confirmed in vivo expression of 10 of 11 selected genes in other volunteers. Of the 531 genes, 139 were identified in at least two volunteers. These 139 genes fell into several functional categories, including biosynthesis and metabolism, regulation, and cellular processes, such as transcription, translation, cell division, DNA replication and repair, and transport. Detection of genes involved in anaerobic and aerobic respiration indicated that H. ducreyi likely encounters both microenvironments within the Pustule. Other genes detected suggest an increase in DNA damage and stress in vivo. Genes involved in virulence in other bacterial pathogens and 32 genes encoding hypothetical proteins were identified, and may represent novel virulence factors. We identified three genes, lspA1, lspA2 and tadA, known to be required for virulence in humans. This is the first study to broadly define transcripts expressed by H. ducreyi in humans.
Diane M. Janowicz - One of the best experts on this subject based on the ideXlab platform.
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Determination of an Interaction Network between an Extracellular Bacterial Pathogen and the Human Host
'American Society for Microbiology', 2019Co-Authors: Brad Griesenauer, Diane M. Janowicz, Kate R Fortney, Tuan M. Tran, Paula Johnson, Hongyu Gao, Stephen Barnes, Landon S. Wilson, Yunlong Liu, Stanley M SpinolaAbstract:Dual RNA sequencing (RNA-seq) offers the promise of determining an interactome at a transcriptional level between a bacterium and the host but has yet to be done on any bacterial infection in human tissue. We performed dual RNA-seq and metabolomics analyses on wounded and infected sites following experimental infection of the arm with H. ducreyi. Our results suggest that H. ducreyi survives in an abscess by utilizing l-ascorbate as an alternative carbon source, possibly taking advantage of host ascorbic acid recycling, and that H. ducreyi also adapts by upregulating genes involved in anaerobic metabolism and inorganic ion and nutrient transport. To our knowledge, this is the first description of an interaction network between a bacterium and the human host at a site of infection.A major gap in understanding infectious diseases is the lack of information about molecular interaction networks between pathogens and the human host. Haemophilus ducreyi causes the genital ulcer disease chancroid in adults and is a leading cause of cutaneous ulcers in children in the tropics. We developed a model in which human volunteers are infected on the upper arm with H. ducreyi until they develop Pustules. To define the H. ducreyi and human interactome, we determined bacterial and host transcriptomic and host metabolomic changes in Pustules. We found that in vivoH. ducreyi transcripts were distinct from those in the inocula, as were host transcripts in Pustule and wounded control sites. Many of the upregulated H. ducreyi genes were found to be involved in ascorbic acid and anaerobic metabolism and inorganic ion/nutrient transport. The top 20 significantly expressed human pathways showed that all were involved in immune responses. We generated a bipartite network for interactions between host and bacterial gene transcription; multiple positively correlated networks contained H. ducreyi genes involved in anaerobic metabolism and host genes involved with the immune response. Metabolomic studies showed that Pustule and wounded samples had different metabolite compositions; the top ion pathway involved ascorbate and aldarate metabolism, which correlated with the H. ducreyi transcriptional response and upregulation of host genes involved in ascorbic acid recycling. These data show that an interactome exists between H. ducreyi and the human host and suggest that H. ducreyi exploits the metabolic niche created by the host immune response
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The Human Skin Microbiome Associates with the Outcome of and Is Influenced by Bacterial Infection
Mbio, 2015Co-Authors: Julia J. Van Rensburg, Sheila Ellinger, Beth Zwickl, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, David E NelsonAbstract:ABSTRACT The influence of the skin microbiota on host susceptibility to infectious agents is largely unexplored. The skin harbors diverse bacterial species that may promote or antagonize the growth of an invading pathogen. We developed a human infection model for Haemophilus ducreyi in which human volunteers are inoculated on the upper arm. After inoculation, papules form and either spontaneously resolve or progress to Pustules. To examine the role of the skin microbiota in the outcome of H. ducreyi infection, we analyzed the microbiomes of four dose-matched pairs of “resolvers” and “Pustule formers” whose inoculation sites were swabbed at multiple time points. Bacteria present on the skin were identified by amplification and pyrosequencing of 16S rRNA genes. Nonmetric multidimensional scaling (NMDS) using Bray-Curtis dissimilarity between the preinfection microbiomes of infected sites showed that sites from the same volunteer clustered together and that Pustule formers segregated from resolvers ( P = 0.001, permutational multivariate analysis of variance [PERMANOVA]), suggesting that the preinfection microbiomes were associated with outcome. NMDS using Bray-Curtis dissimilarity of the endpoint samples showed that the Pustule sites clustered together and were significantly different than the resolved sites ( P = 0.001, PERMANOVA), suggesting that the microbiomes at the endpoint differed between the two groups. In addition to H. ducreyi, Pustule-forming sites had a greater abundance of Proteobacteria , Bacteroidetes , Micrococcus , Corynebacterium , Paracoccus , and Staphylococcus species, whereas resolved sites had higher levels of Actinobacteria and Propionibacterium species. These results suggest that at baseline, resolvers and Pustule formers have distinct skin bacterial communities which change in response to infection and the resultant immune response. IMPORTANCE Human skin is home to a diverse community of microorganisms, collectively known as the skin microbiome. Some resident bacteria are thought to protect the skin from infection by outcompeting pathogens for resources or by priming the immune system9s response to invaders. However, the influence of the skin microbiome on the susceptibility to or protection from infection has not been prospectively evaluated in humans. We characterized the skin microbiome before, during, and after experimental inoculation of the arm with Haemophilus ducreyi in matched volunteers who subsequently resolved the infection or formed abscesses. Our results suggest that the preinfection microbiomes of Pustule formers and resolvers have distinct community structures which change in response to the progression of H. ducreyi infection to abscess formation.
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role played by cd4 foxp3 regulatory t cells in suppression of host responses to haemophilus ducreyi during experimental infection of human volunteers
The Journal of Infectious Diseases, 2010Co-Authors: Klara Tennerracz, Diane M. Janowicz, Kate R Fortney, Barry P. Katz, Paul Racz, Stanley M SpinolaAbstract:Haemophilus ducreyi is the causative agent of the sexually transmitted disease chancroid, a genital ulcer disease that facilitates the acquisition and transmission of HIV-1 [1]. In order to understand the immunopathogenesis of H. ducreyi infection, we developed a human challenge model, in which the skin of the upper arm of healthy adult volunteers is inoculated with the H. ducreyi strain 35000HP (HP, human passaged) or its derivatives [2]. After inoculation, papules form within 24 hours and either spontaneously resolve or evolve into Pustules within 2 to 5 days. Experimental Pustules and natural ulcers are identical histologically and signify immunological failure. Both innate and adaptive immune cells including neutrophils, macrophages, myeloid dendritic cells (DC), NK cells and memory/effector T cells are recruited to experimental Pustules and natural ulcers [3–5]. In Pustules, neutrophils coalesce to form an epidermal abscess, and macrophages form a collar at the base of the abscess. Below the collar, there is a dermal infiltrate of T cells, NK cells and macrophages. Despite this response, H. ducreyi replicates and persists at infected sites. In Pustules and natural ulcers, the bacterium colocalizes with neutrophils and macrophages and remains extracellular [6, 7]. Thus, evasion of phagocytosis is a major mechanism of bacterial survival in experimental and natural infection. Regulatory T (Treg) cells actively suppress the function of the adaptive and innate immune systems [8]. Treg cells are essential for maintaining self-tolerance and immune homeostasis. Treg cells use cell-contact inhibition or soluble factors to control collateral tissue damage mediated by immune responses. However, they also prevent sterilizing antimicrobial immunity and promote pathogen persistence during infection. Two major types of Treg cells have been described based on their origin of generation [9]. Naturally occurring Treg (nTreg) cells are generated in the thymus and express CD25+ and the transcription factor forkhead box P3 (FOXP3), which is critical for their development and function. Treg cells can also be converted from mature CD4+CD25− T cells in peripheral tissues under immunosuppressive conditions, such as exposure to IL-10, transforming growth factor-β (TGF-β) or indoleamine 2,3-dioxygenase (IDO) made by APC [10–14]. These inducible Treg (iTreg) cells are classified as IL-10-producing Tr1 cells, TGF-β-producing Th3 cells and inducible FOXP3+ Treg cells. After infection with pathogens, Treg cells accumulate at infected sites through recruitment, retention, proliferation and/or conversion [15]. Clinical and laboratory data suggest that Treg cells may be involved in the formation of H. ducreyi-induced Pustules. Approximately 16% of volunteers develop hypertrophic scars at H. ducreyi-infected sites that are biopsied [2]. Hypertrophic scar formation is associated with production of TGF-β, which promotes the development of Treg cells. Upon exposure to H. ducreyi, monocyte-derived DC from Pustule formers upregulate transcripts of markers that foster Treg development [16]. Indeed, some H. ducreyi-specific T cell lines and clones isolated from Pustules have characteristics of Treg cells in that they produce IL-10 and IFN-γ [17]. Here, we tested the hypothesis that Treg cells accumulate in H. ducreyi-infected Pustules. We found that CD4+FOXP3+ T cells were enriched in experimental Pustules and were mainly located at the base of Pustules. Most of the CD4+FOXP3+ T cells expressed phenotypic markers characteristic of circulating CD4+FOXP3+ Treg cells. We also showed that the CD4+FOXP3+ T cells were unable to produce effector cytokines and that depletion of CD4+CD25+ T cells from PBMCs of persons who formed Pustules and uninfected donors increased proliferative response of CD4 T cells to H. ducreyi.
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experimental infection of human volunteers with haemophilus ducreyi fifteen years of clinical data and experience
The Journal of Infectious Diseases, 2009Co-Authors: Diane M. Janowicz, Barry P. Katz, Susan Ofner, Stanley M SpinolaAbstract:Haemophilus ducreyi causes chancroid, which facilitates transmission of human immunodeficiency virus type 1. To better understand the biology of H. ducreyi, we developed a human inoculation model. In the present article, we describe clinical outcomes for 267 volunteers who were infected with H. ducreyi. There was a relationship between papule formation and estimated delivered dose. The outcome (either Pustule formation or resolution) of infected sites for a given subject was not independent; the most important determinants of Pustule formation were sex and host effects. When 41 subjects were infected a second time, their outcomes segregated toward their initial outcome, confirming the host effect. Subjects with Pustules developed local symptoms that required withdrawal from the study after a mean of 8.6 days. There were 191 volunteers who had tissue biopsy performed, 173 of whom were available for follow-up analysis; 28 (16.2%) of these developed hypertrophic scars, but the model was otherwise safe. Mutant-parent trials confirmed key features in H. ducreyi pathogenesis, and the model has provided an opportunity to study differential human susceptibility to a bacterial infection.
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identification of haemophilus ducreyi genes expressed during human infection
Microbiology, 2008Co-Authors: Margaret E Bauer, Diane M. Janowicz, Kate R Fortney, Robert S Munson, Alistair Harrison, Stanley M SpinolaAbstract:To identify Haemophilus ducreyi transcripts that are expressed during human infection, we used selective capture of transcribed sequences (SCOTS) with RNA isolated from Pustules obtained from three volunteers infected with H. ducreyi, and with RNA isolated from broth-grown bacteria used to infect volunteers. With SCOTS, competitive hybridization of tissue-derived and broth-derived sequences identifies genes that may be preferentially expressed in vivo. Among the three tissue specimens, we identified 531 genes expressed in vivo. Southern blot analysis of 60 genes from each tissue showed that 87 % of the identified genes hybridized better with cDNA derived from tissue specimens than with cDNA derived from broth-grown bacteria. RT-PCR on nine additional Pustules confirmed in vivo expression of 10 of 11 selected genes in other volunteers. Of the 531 genes, 139 were identified in at least two volunteers. These 139 genes fell into several functional categories, including biosynthesis and metabolism, regulation, and cellular processes, such as transcription, translation, cell division, DNA replication and repair, and transport. Detection of genes involved in anaerobic and aerobic respiration indicated that H. ducreyi likely encounters both microenvironments within the Pustule. Other genes detected suggest an increase in DNA damage and stress in vivo. Genes involved in virulence in other bacterial pathogens and 32 genes encoding hypothetical proteins were identified, and may represent novel virulence factors. We identified three genes, lspA1, lspA2 and tadA, known to be required for virulence in humans. This is the first study to broadly define transcripts expressed by H. ducreyi in humans.
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a case of ceftriaxone induced acute generalized exanthematous pustulosis generalized pustular psoriasis overlap
Case Reports in Dermatology, 2018Co-Authors: Jianfeng Zheng, Xuemei Yi, Yangfeng DingAbstract:Acute generalized exanthematous pustulosis, characterized by subcorneal or superficial intraepidermal Pustules, is induced by drugs in more than 90% of cases. Psoriasis is an autoimmune disease triggered by different conditions in genetically susceptible people. Generalized pustular psoriasis is an acute and severe clinical form of psoriasis, which usually occurs in patients with psoriasis undergoing aggravating factors. In this report the authors have reported a 40-year-old male patient with primary syphilis who developed generalized pustular dermatosis after the use of ceftriaxone. On the third day after ceftriaxone treatment, complete regression of the syphilis lesions was reached. While on the sixth day, erythematous pustular lesions accompanied with fever were observed on the whole body. A personal history of psoriasis and histopathological findings with psoriasiform changes and subcorneal Pustule favored the diagnosis. After discontinuation of ceftriaxone, the patient's condition slowly improved until he had plaque-type psoriasis 3 weeks later. A heterozygous c.115 + 6T>C missense substitution of IL36RN related to the pathogenesis of acute generalized exanthematous pustulosis/generalized pustular psoriasis was identified.
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A Case of Ceftriaxone-Induced Acute Generalized Exanthematous Pustulosis/Generalized Pustular Psoriasis Overlap
Karger Publishers, 2018Co-Authors: Jianfeng Zheng, Yunlu Gao, Yangfeng DingAbstract:Acute generalized exanthematous pustulosis, characterized by subcorneal or superficial intraepidermal Pustules, is induced by drugs in more than 90% of cases. Psoriasis is an autoimmune disease triggered by different conditions in genetically susceptible people. Generalized pustular psoriasis is an acute and severe clinical form of psoriasis, which usually occurs in patients with psoriasis undergoing aggravating factors. In this report the authors have reported a 40-year-old male patient with primary syphilis who developed generalized pustular dermatosis after the use of ceftriaxone. On the third day after ceftriaxone treatment, complete regression of the syphilis lesions was reached. While on the sixth day, erythematous pustular lesions accompanied with fever were observed on the whole body. A personal history of psoriasis and histopathological findings with psoriasiform changes and subcorneal Pustule favored the diagnosis. After discontinuation of ceftriaxone, the patient’s condition slowly improved until he had plaque-type psoriasis 3 weeks later. A heterozygous c.115 + 6T>C missense substitution of IL36RN related to the pathogenesis of acute generalized exanthematous pustulosis/generalized pustular psoriasis was identified
